1

What is compartmentalization?

• Mainly in eukaryotic cells.

• Biochemical reactions take place in an organized way in cells (different reactions with different requirements

at the same time)→ these reactions have to take place in a spatially separated (compartmentalized) way

• cell nucleus

• mitochondria

• endoplasmic reticulum

• Golgi

• secretory vesicles

• endosomes

• lysosomes

• peroxisomes

• cytoskeleton

• centrosome

• ribosome

• nucleolus

• Cajal bodies, …

Organelles: specialized structures in cells that have

specific functions (less restrictive definition)

membrane-bounded

organelles: according

to the generally

accepted, more

restrictive definition

only these are

considered to be

organelles, i.e.

specialized structures

having specific

function bounded by

a membrane

2

What is the advantage and disadvantage of compartmentalization?

Advantage of compartmentalization:

• membrane-bound organelles establish special conditions favoring a given set of biological functions

(e.g. lysosome – hydrolytic degradation, mitochondrion – ATP generation, etc.)

• different reactions with different requirements at the same time

Disadvantage of compartmentalization:

• transport across compartment boundaries is complex and often energy requiring

What are the realizations of compartmentalization?

1. „Virtual compartments” (without membranes):

2. Compartmentalization (by membranes, only in eukaryotes) →

transmembrane transport is necessary which is often energy-

consuming.

B) Droplet organelles (generated by liquid-liquid phase separation – liquid

droplets) → diffusion takes place inside them and they exchange

material with their surroundings (e.g. nucleolus, Cajal bodies, etc.)

A) Enzyme complexes (in eukaryotes and prokaryotes) → efficient since

there is no need for diffusion (e.g. mammalian fatty acid synthase)

3

What is endosymbiosis? Which compartments were generated by endosymbiosis?

• eukaryotic organism, already equipped with adequate

cytoskeleton and internal membrane systems

• phagocytosed a prokaryotic organism (engulfed)

• they started to live together for their mutual benefit.

• Mitochondria came about when an aerobic prokaryote

performing oxidative phosphorylation was internalized.

The origin of peroxisomes is similar.

What are the proofs for the bacterial origin of mitochondria?

cytosolnucleus

extracellular space

vesicleslysosome

ER, nuclear membrane

mitochondrion

peroxisome (?)

Golgi

1. DNA

• circular DNA

• no introns

• polycistronic mRNA

• The genetic code of mitochondria is different from the universal

code in some cases.

2. Ribosomes resembling prokaryotic ribosomes

3. Size of mitochondria (similar to the size of bacteria (~ 1 μm)).

4. Two membranes (the composition of the inner membrane is similar

to that of bacterial membranes).

5. Mitochondria divide independently of the host cells by binary

fission, like bacteria.

What about the peroxisome?

4

How can you classify the compartments from the stand point of intracellular transport?

cytosol

extracellular space

vesicleslysosome

ER, nuclear membrane

mitochondrion

peroxisome

nucleus

Golgi

1. endomembrane system: ER, Golgi, lysosome,

vesicles (endo- and exocytic, transport) and the

lumen of the nuclear membrane (+EC space,

plasma membrane)

2. cytoplasmic compartment:

2a. cytosol

2b. organelles communicating with the

cytosol by non-vesicular transport: nucleus,

peroxisome and the mitochondrion

transport between individual

organelles by means of vesicular

transport

diffusion/directed motion +

transmembrane transport

5

What is the distance which can be covered efficiently (in a couple of seconds) by diffusion in a

living organ? Why?

• diffusion is an efficient way to transport molecules up to ∼∼∼∼100 µµµµm

• it is not efficient for transport to larger distances

• mean squared displacement:

2 2 2 26r x y z Dt∆ = ∆ + ∆ + ∆ = average displacement t∼

= ⇒ ∼ ∼3

1 1kTD D

f f MW

k – Boltzmann constant

T – absolute temperature

f – form factor

MW – molecular weight

glycine

(MW=75)

glucose

(MW=180)

ordinary (40

kDa) protein

D (µm2/s) 103 5�102 102

distance travelledtime of diffusion (sec)

protein glucose glycine

1 µm 0.0017 0.00033 0.00017

10 µm (size of a

eukaryotic cell)0.17 0.033 0.017

100 µm (max. distance

of cells from capillaries)16.7 3.3 1.7

1 mm 1667 333 166.7

1 cm 166667 33333 16667

weak dependence on MW

6

Why does the diffusion rate of proteins decline steeply with their molecular weight?

The rate of diffusion is decreased by:

• molecular weight dependent, static filtering effect of the cytosolic matrix with a pore size of ~50 nm

• dynamic filtering effect of macromolecules

• specific interaction of proteins with the cytoskeleton (or DNA in the nucleus)

7

What are the main functions of the smooth endoplasmic reticulum (SER)? Which organs

contain a large amount of it?

• SER belongs to the endomembrane system

• storage and release of Ca2+ (heart, skeletal muscles → sarcoplasmic reEculum)

• biochemical reactions: synthesis of lipids, cholesterol, steroid hormones (adrenal gland),

gluconeogenesis (liver, glucose-6-phosphate→glucose)

• detoxification (liver)

What is detoxification?

� A lot of toxic compounds are hydrophobic, and bind to serum proteins in the blood.

� Protein-bound molecules cannot leave the circulation through the kidneys.

� The liver converts most of these hydrophobic compounds to hydrophilic, water-soluble molecules by

• mono-oxidation (hydroxylation) of the toxin (phase I reaction) → by cytochrome P450 enzyme

• followed by conjugation of hydrophilic groups (e.g. glucuronic acid) to it (phase II reaction)

� These conjugated molecules can leave the body through the liver and bile ducts.

Mutagenic, carcinogenic substances can also be produced by hydroxilation.

8

What is the main function of the lysosomes?

It belongs to the endomembrane system.

Function:

Hydrolytic degradation of substances.

Lysosomal enzymes are synthesized in the rough ER.

Role of acidic pH:

• denatures substances

• acid hydrolases work optimally at acidic pH generated

by H+-ATPases located in the membrane

Storage diseases:

• due to a congenital lack of a certain type of lysosomal

hydrolase the substrates of the enzyme accumulate

causing major neurological and hematological symptoms

• example: Tay-Sachs disease caused by a mutation of

hexosaminidase A, a ganglioside-degrading enzyme.

What is the difference between primary and secondary lysosomes?

A primary lysosome contains only the enzymes, but not the materials to be digested.

Secondary lysosomes contain both the acid hydrolases and the materials to be degraded.

9

How can materials be transported into the lysosomes?

Endocytosis/phagocytosis

Autophagy:

• A cell decomposes its own

components in its lysosomes.

• An organelle or cellular constituent

separated from the rest of the cell by a

membrane is an autophagosome which

is converted to an autolysosome by

fusion with a lysosome.

10

What are the different mechanisms of autophagy?

Autophagy takes place to replenish food stores in starvation and to get rid of worn-out organelles and proteins.

3. Chaperone-mediated autophagy

1. Macroautophagy (usually

simply autophagy)

LAMP-2A

cytosolic

protein

hsc70 and cochaperons

lysosomal hsc70

2. Microautophagy – lysosomes

directly wrap around cytosolic

constituents and ingest them

11

What is the main function of the peroxisome?

A membrane-bound organelle, which may have an endosymbiotic origin and it is present in large amount in

liver and kidney cells. Peroxisomal enzymes are synthesized in the cytoplasm.

Function:

• Oxidation of fatty acids, alcohol, amino acids by peroxisomal oxidases (e.g. peroxidases) → H2O2 is formed

• Oxidation of D-amino acids and very long chain fatty acids is exclusively carried out in peroxisomes

• Neutralization of hydrogen-peroxide by catalase → 2 H2O2 → 2 H2O + O2

Organelle Function Explanation, differences between organelles

mitochondrionoxidative degradation,

ATP generation

1. oxidation is carried out in multiple steps (electron

transport chain), energy is released in a controlled

way like in a nuclear power plant → energy can be

stored (electricity in a power plant, ATP in

mitochondria)

2. the final electron acceptor (O2) is converted to H2O

lysosome hydrolytic degradation

peroxisomeoxidative degradation, no

ATP generation

1. peroxisomal oxidation is a single step reaction →energy is released as heat → no energy is stored

similar to the explosion of a nuclear bomb

2. the final electron acceptor (O2) is converted to H2O2

12

ER signal sequence:

A sequence composed of ~15-30, mainly hydrophobic, amino acids on the N-terminal of certain

polypeptides. It is necessary and sufficient to induce transport of a protein into the ER.

How does the ribosome know whether to bind to the ER?

13

The ER signal sequence binds to the SRP (signal recognition particle) in the cytoplasm and directs the

ribosome to the docking site (signal recognition particle receptor) on the surface of the ER. In the case of

soluble proteins, the signal sequence is removed by the signal peptidase located in the ER lumen.

(sec61)

translation stops

temporarily translation starts

again

How can a protein get into the ER?

14

Besides the ER signal sequence what other special sequence do integral

membrane proteins have?

15

What is quality control in the ER?

• Ensures that only correctly folded proteins, whose appropriate 3D structure has formed spontaneously or

with the help of chaperon proteins (e.g. Bip), can leave the ER.

• If the faulty conformation of a proteins cannot be corrected, the protein will get degraded by proteasomes.

Hsp70 (Bip) binds to

hydrophobic patches

on the protein to

prevent its

aggregation

incorrect

folding

proteasomal

degradation

correct

folding

transport to the Golgi organelle

16

What are ER-resident proteins?

These proteins are retained in the ER after

folding. The classical ER retention signal is the C-

terminal KDEL sequence (K-Lysine, D-Aspartic

acid, E-Glutamic acid, L-Leucine). This signal

allows for retrieval from the Golgi by KDEL

receptors.

Purpose of bidirectional transport:

1. To maintain the equilibrium

between membranes

2. Retrograde transport of proteins

accidentally / non-specifically

transported

Purpose of bidirectional transport:

1. To maintain the equilibrium

between membranes

2. Retrograde transport of proteins

accidentally / non-specifically

transported

17

QUIZ

1) It carries out hydrolysis.

2) In muscle cells we call it sarcoplasmic reticulum.

3) It is involved in the break down of H2O2.

4) It may have evolved by endosymbiosis.

5) It is involved in the removal of toxic substances in the liver cells.

6) It contains a large amount of catalase.

7) Ribosomes are bound to its outer surface.

8) It carries out oxidation.

9) It functions as a Ca2+ storage.

10) It contains enzymes involved in the post-translational

modification of proteins.

11) Oxidative processes take place inside it and the energy released

is converted to heat.

Smooth-ER (SER), Rough-ER (RER), Golgi, lysosome, peroxisome, mitochondria

1) Lysosome

2) SER

3) Peroxisome

4) Mitochondria, peroxisome

5) SER

6) Peroxisome

7) RER

8) Peroxisome, mitochondria

9) SER

10) RER, Golgi

11) Peroxisome

I. …………………….. is a process used for degrading the cells own organelle.

II. …………… and …………….. can be oxidized exclusively in peroxisomes.

I. Autophagy

II. D-amino acids AND

long chain fatty acids